| 1. |
- Acheva, Anna, et al.
(author)
-
Presence of Stromal Cells Enhances Epithelial-to-Mesenchymal Transition (EMT) Induction in Lung Bronchial Epithelium after Protracted Exposure to Oxidative Stress of Gamma Radiation
- 2019
-
In: Oxidative Medicine and Cellular Longevity. - : Hindawi Limited. - 1942-0900 .- 1942-0994. ; 2019
-
Journal article (peer-reviewed)abstract
- The aim of the study was to investigate the role of a microenvironment in the induction of epithelial-to-mesenchymal transition (EMT) as a sign of early stages of carcinogenesis in human lung epithelial cell lines after protracted low-dose rate gamma-radiation exposures. BEAS-2B and HBEC-3KT lung cell lines were irradiated with low-dose rate gamma-rays (Cs-137, 1.4 or 14 mGy/h) to 0.1 or 1 Gy with or without adding TGF-beta. TGF-beta-treated samples were applied as positive EMT controls and tested in parallel to find out if the radiation has a potentiating effect on the EMT induction. To evaluate the effect of the stromal component, the epithelial cells were irradiated in cocultures with stromal MRC-9 lung fibroblasts. On day 3 post treatment, the EMT markers: alpha-SMA, vimentin, fibronectin, and E-cadherin, were analyzed. The oxidative stress levels were evaluated by 8-oxo-dG analysis in both epithelial and fibroblast cells. The protracted exposure to low Linear Energy Transfer (LET) radiation at the total absorbed dose of 1 Gy was able to induce changes suggestive of EMT. The results show that the presence of the stromal component and its signaling (TGF-beta) in the cocultures enhances the EMT. Radiation had a minor cumulative effect on the TGF-beta-induced EMT with both doses. The oxidative stress levels were higher than the background in both epithelial and stromal cells post chronic irradiation (0.1 and 1 Gy); as for the BEAS-2B cell line, the increase was statistically significant. We suggest that the induction of EMT in bronchial epithelial cells by radiation requires more than single acute exposure and the presence of stromal component might enhance the effect through free radical production and accumulation.
|
|
| 2. |
- Cheng, Lei, et al.
(author)
-
Comet assay reveals an interaction of DNA lesions and impairment of DNA repair in peripheral blood lymphocytes simultaneously exposed to alpha particles and X-rays
-
Other publication (other academic/artistic)abstract
- The biological effectiveness of ionising radiation is related to the ionisation density which is defined by the linear energy transfer LET. Radiation quality factors are applied to calculate the equivalent dose in the field of radiation protection and the biologically effective dose in the field of radiotherapy. Additivity is assumed in exposure scenarios where radiations of different qualities are mixed. We have carried out a series of studies on the cytogenetic effect of exposing human peripheral blood lymphocytes to a mixed beam of the high LET alpha radiation and low LET X-rays and could demonstrate that both radiations interact in producing more chromosomal aberrations than expected based on additivity. The aim of the present investigation was to look at the mechanism of the interaction, especially with respect to the question if it is due to an augmented level of initial damage or impaired DNA repair. The level of DNA damage and the kinetics of damage repair was quantified by the alkaline comet assay. The levels of phosphorylated, key DNA damage response (DDR) proteins were also measured by Western blotting. The results revealed that alpha particles and X-rays interact in inducing DNA damage above the level predicted by assuming additivity and that the repair of damage occurs with a delay. Moreover, the activation levels of the key DDR proteins ATM, p53 and DNA PK were highest in cells exposed to mixed beams substantiating the idea exposure to mixed beams presents a challenge to the cellular DNA damage response system.
|
|
| 3. |
- Cheng, Lei, et al.
(author)
-
Modulation of radiation-induced cytogenetic damage in human peripheral blood lymphocytes by hypothermia
- 2015
-
In: Mutation research. Genetic toxicology and environmental mutagenesis. - : Elsevier BV. - 1383-5718 .- 1879-3592. ; 793:SI, s. 96-100
-
Journal article (peer-reviewed)abstract
- Purpose: Recent studies have shown that low temperature (hypothermia) at exposure can act in a radioprotective manner at the level of cytogenetic damage. The mechanisms of this phenomenon are not understood, but it was suggested to be due to hypothermia-induced perturbations of the cell cycle. The purpose of the present study was to detect whether a reduced frequency of micronuclei is observed in peripheral blood lymphocytes (PBL) irradiated at low temperature and harvested sequentially at 3 time points. Additionally, the level of apoptosis was estimated by microscopic analysis of the MN slides. Materials and methods: Experiments were carried out with blood drawn from three donors at the Stockholm University and from three donors at the Jan Kochanowski University. Prior to irradiation, blood samples were incubated for 20 mm and irradiated at the respective temperature (0 degrees C and 37 degrees C) with gamma rays. Whole blood cultures were set up, cytochalasin B was added after 44h of irradiation and the samples were harvested after 72,96 and 120 h of incubation time. Results and conclusions: The frequency of micronuclei was markedly lower in PBL harvested at 72h, 96 h and 120 h following irradiation at 0 degrees C as compared to 37 degrees C. This indicates that the temperature effect observed in peripheral blood lymphocytes after irradiation is not related to a temporary perturbation of the cell cycle. Also, it is not due to selective elimination of damaged cells by apoptosis.
|
|
| 4. |
- Cheng, Lei, et al.
(author)
-
Simultaneous induction of dispersed and clustered DNA lesions compromises DNA damage response in human peripheral blood lymphocytes
- 2018
-
In: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 13:10
-
Journal article (peer-reviewed)abstract
- Due to its ability to induce DNA damage in a space and time controlled manner, ionising radiation is a unique tool for studying the mechanisms of DNA repair. The biological effectiveness of ionising radiation is related to the ionisation density which is defined by the linear energy transfer (LET). Alpha particles are characterised by high LET, while X-rays by low LET values. An interesting question is how cells react when exposed to a mixed beam of high and low LET radiation. In an earlier study carried out with human peripheral blood lymphocytes (PBL) we could demonstrate that alpha radiation X-rays interact in producing more chromosomal aberrations than expected based on additivity. The aim of the present investigation was to look at the mechanism of the interaction, especially with respect to the question if it is due to an augmented level of initial damage or impaired DNA repair. PBL were exposed to various doses of alpha particles, X-rays and mixed beams. DNA damage and the kinetics of damage repair was quantified by the alkaline comet assay. The levels of phosphorylated, key DNA damage response (DDR) proteins ATM, p53 and DNA-PK were measured by Western blotting and mRNA levels of 6 damage-responsive genes were measured by qPCR. Alpha particles and X-rays interact in inducing DNA damage above the level predicted by assuming additivity and that the repair of damage occurs with a delay. The activation levels of DDR proteins and mRNA levels of the studied genes were highest in cells exposed to mixed beams. The results substantiate the idea that exposure to mixed beams presents a challenge for the cellular DDR system.
|
|
| 5. |
- Dang, Li, et al.
(author)
-
Radioprotective effect of hypothermia on cells - a multiparametric approach to delineate the mechanisms
- 2012
-
In: International Journal of Radiation Biology. - : Informa UK Limited. - 0955-3002 .- 1362-3095. ; 88:7, s. 507-514
-
Journal article (peer-reviewed)abstract
- Purpose: Low temperature (hypothermia) during irradiation of cells has been reported to have a radioprotective effect. The mechanisms are not fully understood. This study further investigates the possible mechanisms behind hypothermia-mediated radioprotection. Materials and methods: Human lymphoblastoid TK6 cells were incubated for 20 min at 0.8 or 37 degrees C and subsequently exposed to 1 Gy of gamma- or X-rays. The influence of ataxia telangiectasia mutated (ATM)-mediated double-strand break signalling and histone deacetylase-dependent chromatin condensation was investigated using the micronucleus assay. Furthermore, the effect of hypothermia was investigated at the level of phosphorylated histone 2AX (gamma H2AX) foci, clonogenic cell survival and micronuclei in sequentially-harvested cells. Results: The radioprotective effect of hypothermia (called the temperature effect [TE]) was evident only at the level of micronuclei at a single fixation time, was not influenced by the inhibition of ATM kinase activity and completely abolished by the histone deacetylase inhibition. No TE was seen at the level of gamma H2AX foci and cell survival. Conclusions: We suggest that low temperature during irradiation can induce a temporary cell cycle shift, which could lead to a reduced micronucleus frequency. Future experiments focused on cell cycle progression are needed to confirm this hypothesis.
|
|
| 6. |
- Danielsson, Daniel, et al.
(author)
-
Influence of genetic background and oxidative stress response on risk of mandibular osteoradionecrosis after radiotherapy of head and neck cancer
- 2016
-
In: Head and Neck. - : Wiley. - 1043-3074 .- 1097-0347. ; 38:3, s. 387-393
-
Journal article (peer-reviewed)abstract
- Background: Osteoradionecrosis (ORN) of the mandible is a severe complication of head and neck radiotherapy (RT) treatment, where the impact of individual radiosensitivity has been a suggested explanation. Methods: A cohort of patients with stage II/III ORN was compared to matched controls. Blood was collected and irradiated in vitro to study the capacity to handle radiation-induced oxidative stress. Patients were also genotyped for 8 single-nucleotide polymorphisms (SNPs) in genes involved in the oxidative stress response. Results: A difference in 8-oxo-7,8-dihydro-2-deoxyguanosine (8-oxo-dG) levels was found between the patient cohorts (p = 0.01). The SNP rs1695 in glutathione s-transferase p1 (GSTP1) was also found to be more frequent in the patients with ORN (p = .02). Multivariate analysis of the clinical and biological factors revealed concomitant brachytherapy plus the 2 biomarkers to be significant factors which influense risk of mandibular osteoradionecrosis after radiotherapy of head and neck cancer. Conclusion: The current study indicates that oxidative stress response contributes to individual radiosensitivity and healthy tissue damage caused by RT and may be predicted by biomarker analysis.
|
|
| 7. |
- Gałecki, Maciej, et al.
(author)
-
Precision of scoring radiation-induced chromosomal aberrations and micronuclei by unexperienced scorers
- 2019
-
In: International Journal of Radiation Biology. - : Informa UK Limited. - 0955-3002 .- 1362-3095. ; 95:9, s. 1251-1258
-
Journal article (peer-reviewed)abstract
- Purpose: Dose assessment plays an important role in case of radiological accidents and can be performed by scoring structural changes of chromosome morphology induced in cells by ionizing radiation. The results of such a test are biased by scorer experience, therefore, simple to learn assays are recommended to be used when fast analysis of a large amount of data is needed. The aim of this study was to compare the performance of two radiobiological assays - chromosomal aberrations and micronuclei - by unexperienced scorers with the reference values generated by an expert.Materials and methods: Each participant of an EU-funded two-week radiobiology course was asked to score Chinese hamster ovary cells exposed to gamma radiation up to 4 Gy. The congruence of students' and expert's scores at each dose and the coherence of the dose-response curve parameters between the students were investigated.Results: Micronucleus test tended to be faster and easier to learn than scoring chromosomal aberrations. However, both assays carried out by inexperienced students showed reasonable dose-response curves.Conclusions: In the case of a large radiological accident involving many casualties, the unexperienced scorers would support the process of biodosimetric triage by cytogenetic biological dosimetry.
|
|
| 8. |
- Lisowska, Halina, et al.
(author)
-
Hypothermia modulates the DNA damage response to ionizing radiation in human peripheral blood lymphocytes
- 2018
-
In: International Journal of Radiation Biology. - : Informa UK Limited. - 0955-3002 .- 1362-3095. ; 94:6, s. 551-557
-
Journal article (peer-reviewed)abstract
- Purpose: Low temperature at exposure has been shown to act in a radioprotective manner at the level of cytogenetic damage. It was suggested to be due to an effective transformation of DNA damage to chromosomal damage at low temperature. The purpose of the study was to analyze the kinetics of aberration formation during the first hours after exposing human peripheral blood lymphocytes to ionizing radiation at 0.8 degrees C and 37 degrees C.Materials and methods: To this end, we applied the technique of premature chromosome condensation. In addition, DNA damage response was analyzed by measuring the levels of phosphorylated DNA damage responsive proteins ATM, DNA-PK and p53 and mRNA levels of the radiation-responsive genes BBC3, FDXR, GADD45A, XPC, MDM2 and CDKN1A.Results: A consistently lower frequency of chromosomal breaks was observed in cells exposed at 0.8 degrees C as compared to 37 degrees C already after 30minutes postexposure. This effect was accompanied by elevated levels of phosphorylated ATM and DNA-PK proteins and a reduced immediate level of phosphorylated p53 and of the responsive genes.Conclusions: Low temperature at exposure appears to promote DNA repair leading to reduced transformation of DNA damage to chromosomal aberrations.
|
|
| 9. |
|
|
| 10. |
- Sollazzo, Alice, 1981-, et al.
(author)
-
Alpha particles and X-rays interact in inducing DNA damage in U2OS cells
- 2017
-
In: Radiation Research. - 0033-7587 .- 1938-5404. ; 188:4, s. 400-411
-
Journal article (peer-reviewed)abstract
- The survivors of atomic bomb explosions in Hiroshima and Nagasaki are monitored for health effect within the Life Span Study (LSS). The LSS results represent the most important source of knowledge about cancer effects of ionizing radiation and they form the basis for the radiation protection system. One uncertainty connected to deriving universal risk factors from these results is related to the problem of mixed radiation qualities. The atomic bomb explosions generated a mixed beam of the sparsely ionizing gamma radiation and densely ionizing neutrons and what is not taken into consideration is the problem of a possible interaction of the two radiation types in inducing biological effects. The existence of such interaction would suggest that the application of risk factors derived from the LSS to predict cancer effects after exposure to pure gamma radiation (such as in the Fukushima prefecture) leads to an overestimation of risk.In order to analyze the possible interaction of radiation types a mixed beam exposure facility was constructed where cells can be exposed to sparsely ionizing X-rays and densely ionizing alpha particles. U2OS cells were used, which are stably transfected with a plasmid coding for the DNA repair gene 53BP1 coupled to a gene coding for the green fluorescent protein GFP. Induction and repair of DNA damage which are known to be related to cancer induction were analyzed. The results suggest that alpha particles and X-rays interact, leading to cellular, and possibly cancer effects not predictable based on assuming simple additivity of the individual mixed beam components.
|
|
| 11. |
- Sollazzo, Alice, 1981-
(author)
-
Cellular responses to combined irradiation with alpha particles and X-rays
- 2017
-
Doctoral thesis (other academic/artistic)abstract
- Mixed radiation fields, where different ionizing particles act together, are very important in radiobiology and in radiation protection. Mixed beams are not only the most common form of radiation exposure, but the prediction of their biological effect is also full of uncertainties. Currently, prediction of the biological damage of exposure to mixed radiation fields is based on the default assumption of simple additivity between the effects of all the radiation in the field. This assumption has been proven to be incorrect. Indeed, the simultaneous effect of different radiation qualities has been shown to be greater than additive, namely synergistic. This implicates that, for instance, the predicted cancer risk for astronauts, that remain a prolonged time in space, is currently underestimated as well as the risk of developing secondary cancer for radiotherapy patients.This thesis aims at understanding the mechanisms behind the cellular response to simultaneous exposure to alpha particles and X-rays (that is referred as mixed beam).Paper I describes the cell killing and the mutagenic effect of mixed beam exposure in human lymphoblastoid wild type and in cells with impaired capacity to repair oxidative DNA damage .We found that oxidative DNA damage plays an important role in the lethal, synergistic effect of mixed beams.Paper II and III investigates whether mixed beams exposure leads to an augmented DNA double strand breaks (DSB) induction or to an altered response of the cellular DSB repair machinery. We found that mixed irradiation resulted in synergistic induction of DSB, and that those lesions were repaired with slow kinetics.Paper IV focuses on the effect of mixed beams at the level of DNA damage in normal cells. Induction and repair of DNA lesions such as DSB, single strand breaks and apurinic sites was quantified using the alkaline comet assay. We found that alpha particles and X-rays interacted in inducing DNA damage. Moreover, although mixed beam exposure resulted in strong activation of the DNA damage response, it resulted in delayed repair.Although more research is needed to fully elucidate the mechanisms behind the detected synergistic effects, our results strongly suggest that an overwhelmed DNA-repair system causes delay in repair of damage.
|
|
| 12. |
- Sollazzo, Alice, et al.
(author)
-
Interaction of low and high LET radiation in TK6 cells-mechanistic aspects and significance for radiation protection
- 2016
-
In: Journal of Radiological Protection. - : IOP Publishing. - 0952-4746 .- 1361-6498. ; 36:4, s. 721-735
-
Journal article (peer-reviewed)abstract
- Most environmental, occupational and medical exposures to ionising radiation are associated with a simultaneous action of different radiation types. An open question remains whether radiations of different qualities interact with each other to yield effects stronger than expected based on the assumption of additivity. It is possible that DNA damage induced by high linear energy transfer (LET) radiation will lead to an opening of the chromatin structure making the DNA more susceptible to attack by reactive oxygen species (ROS) generated by the low LET radiation. In such case, the effect of mixed beams should be strongly expressed in cells that are sensitive to ROS. The present investigation was carried out to test if cells with an impaired capacity to handle oxidative stress are particularly sensitive to the effect of mixed beams of alpha particles and x-rays. Clonogenic cell survival curves and mutant frequencies were analysed in TK6 wild type (wt) cells and in TK6 cells with a knocked down hMYH glycosylase. The results showed a synergistic effect of mixed beams on clonogenic cell survival of TK6(wt) but not TK6(MYH)-cells. The frequencies of mutants showed a high degree of interexperimental variability without any indications for synergistic effects of mixed beams. TK6(MYH)-cells were generally more tolerant to radiation exposure with respect to clonogenic cell survival but showed a strong increase in mutant frequency. The results demonstrate that exposure of wt cells to a mixed beam of alpha particles and x-rays leads to a detrimental effect which is stronger than expected based on the assumption of additivity. The role of oxidative stress in the reaction of cells to mixed beams remains unclear.
|
|
| 13. |
- Sollazzo, Alice, 1981-, et al.
(author)
-
Live dynamics of 53BP1 DNA damage foci induced by a combination of clustered and dispersed double strand breaks
-
Other publication (other academic/artistic)abstract
- Cells react differently to alpha particle-induced clustered DNA damage and X-ray induced dispersed DNA damage. Little is known about how they cope when both types of damage are induced simultaneously. We used live cell microscopy to analyse the formation and motion of the DNA double strand break (DSB) repair factor 53BP1-GFP foci in U2OS cells exposed to a mixed beam of alpha particles and X-rays. We observed that the kinetics of appearance and decline of mixed beam-induced foci resemble that of after alpha particle irradiation. They showed a fast increase and almost no repair during the imaging time, while X-ray-exposed cells showed a strong increase and decline over the observation interval. Secondly, focus sizes were similar to X-ray-induced foci, with both being significantly smaller than those induced by alpha particles; thirdly, they showed a high mean pixel intensity already at early time point after irradiation. Finally, focus mobility was reduced relative to alpha particle and X-ray-induced foci, which was correlated with a delayed appearance of a fraction of mixed beam-induced foci. These results suggest that cells react to a simultaneous induction of clustered and dispersed DNA damage by sequestering the 53BP1 protein in selected foci, possibly at sites of clustered DNA damage. This happens at the cost of foci forming at chromatin areas containing dispersed, simple damage. Our results highlight that the DNA damage response to a combination of dispersed and clustered DNA damage in the same cell differs significantly from the response to exposure with single radiation types.
|
|
| 14. |
- Sollazzo, Alice, et al.
(author)
-
Live Dynamics of 53BP1 Foci Following Simultaneous Induction of Clustered and Dispersed DNA Damage in U2OS Cells
- 2018
-
In: International Journal of Molecular Sciences. - : MDPI AG. - 1661-6596 .- 1422-0067. ; 19:2
-
Journal article (peer-reviewed)abstract
- Cells react differently to clustered and dispersed DNA double strand breaks (DSB). Little is known about the initial reaction to simultaneous induction of DSBs with different complexities. Here, we used live cell microscopy to analyse the behaviour of 53BP1-GFP (green fluorescence protein) foci formation at DSBs induced in U2OS cells by alpha particles, X-rays or mixed beams over a 75 min period post irradiation. X-ray-induced foci rapidly increased and declined over the observation interval. After an initial increase, mixed beam-induced foci remained at a constant level over the observation interval, similarly as alpha-induced foci. The average areas of radiation-induced foci were similar for mixed beams and X-rays, being significantly smaller than those induced by alpha particles. Pixel intensities were highest for mixed beam-induced foci and showed the lowest level of variability over time as compared to foci induced by alphas and X-rays alone. Finally, mixed beam-exposed foci showed the lowest level of mobility as compared to alpha and X-ray exposure. The results suggest paralysation of chromatin around foci containing clustered DNA damage.
|
|
| 15. |
- Sun, Jiao, et al.
(author)
-
Serum 8-hydroxy-2'-deoxyguanosine (8-oxo-dG) levels are elevated in diabetes patients
- 2015
-
In: International Journal of Diabetes in Developing Countries. - : Springer Science and Business Media LLC. - 0973-3930 .- 1998-3832. ; 35:3, s. 368-373
-
Journal article (peer-reviewed)abstract
- The increased oxidative stress in diabetes is known to contribute to the development of diabetes. We investigate whether serum 8-hydro-2'-deoxyguanosine (8-oxo-dG) is associated with diabetes at the time of first diagnosis and evaluate whether it can be used as a reliable biomarker for the oxidative stress in diabetes. The study was designed as a case control study with two groups: patient with diabetes and control. The diabetes group consisted of a total of 28 patients consulting the hospital for the first time and definitely diagnosed for diabetes, and the control group was composed of 65 healthy subjects. Serum 8-oxo-dG was measured by a competitive enzyme-linked immunosorbent assay (ELISA) kit, specially developed to minimize cross-reaction of 8-oxo-dG antibody with serum guanosine. The average serum 8-oxo-dG levels in patients with diabetes and controls were 0.72 +/- 0.41 and 0.24 +/- 0.14 ng/mL, respectively, statistically significant (p < 0.001). The 8-oxo-dG value was significantly higher in women with diabetes, compared with men with diabetes (p = 0.028). The sensitivity and the specificity of the 8-oxo-dG ELISA assay were 0.80 and 0.96, respectively, and the ROC value was 0.93. This study suggests that increased oxidative stress has an important role in the pathogenesis of diabetes. Serum 8-oxo-dG may be a useful clinical biomarker for the early diagnosis of stress-related diseases, e.g. diabetes and its management.
|
|
